建立人急性粒细胞白血病M_2裸鼠模型

Establishment of human acute myeloblastic Leukemia M_2 type nude mouse model

背景:人实体瘤细胞容易在小鼠体内成瘤,但人荷瘤白血病模型很难构建。通过放射线或环磷酰胺抑制裸鼠免疫系统预处理,可构建低成本、高稳定性的裸鼠模型。目的:探讨融合基因AML/ETO阳性的人急性粒细胞白血病M2白血病细胞Kasumi-1在BALB/c裸鼠体内建立白血病模型的方法。方法:将BALB/c裸鼠以抽签法随机分3组:环磷酰胺组腹腔注射环磷酰胺连续2d后,尾静脉注射Kasumi-1细胞8×105只;照射组给予X射线全身照射,照射当天尾静脉注射Kasumi-1细胞;无预处理组未作任何处理,尾静脉注射Kasumi-1白血病细胞。另取3只正常BALB/c裸鼠作为正常对照。检测外周血涂片、骨髓涂片,流式细胞仪检测骨髓细胞免疫分型,RT-PCR检测白血病细胞瘤负荷,FISH检测骨髓细胞AML/ETO融合基因阳性细胞百分比。结果与结论:未经任何预处理裸鼠建模14d血涂片中白血病细胞达3.5%,骨髓中肿瘤细胞百分比可达40%以上,与FISH和流式细胞仪检测白血病细胞比例一致,且随着接种时间延长,瘤负荷不断增加。全身照射和环磷酰胺注射后的裸鼠瘤负荷高于无预处理组,但仍可带瘤生存60d。正常裸鼠外周血单个核细胞RT-PCR未发现有融和基因AML/ETO,其他3组均可见融和基因AML/ETO的mRNA表达。提示给予环磷酰胺组和X线照射预处理或单纯尾静脉接种Kasimi-1细胞均可建立急性粒细胞白血病M2裸鼠模型。

BACKGROUND： It is easy to established human solid tumor nude mouse model, but for leukemia which is difficult. We inhibited immune system further by radioactive ray or CTX, to decrease cost and increase the stability. OBJECTIVE： To establish a human acute myeloblastic leukemia M2 Kasumi-1 models containing AML/ETO positive genes in BALB/c nude mouse. METHODS： Nude mice were randomly divided into three groups： CTX group was injected CTX 2 mg/day in abdominal cavity for two days, and injected 8×105/mouse Kasumi-1 cells in caudal vein next day; irradiation group was exposed to total body irradiation, and injected 8×105/mouse Kasumi-1 cells in caudal vein that day; untreated group was inoculated with 8×105/mouse Kasumi-1 cells by caudal vein injection. Three additional mice were considered as the normal control group. The blood smearing and bone morrow slides were detected, immunity type of BMC was detected using flow cytometry, loading of leukemic cellular tumor was detected using RT-PCR, and positive ratio of AML/ETO fusion gene was detected using FISH method. RESULTS AND CONCLUSION： After inoculated into untreated nude mice by caudal vein injection for 14 days, the ratio of leukemia cell in blood smearing was 3.5%, and over 40% in bone marrow slides, which was equal to the results of FISH and FCM. The increasing of tumor loading was time-dependent. For irradiation group and CTX treated group, the tumor loading was higher that untreated group, and the cells also survived more than 60 days. AML/ETO band was observed by RT-PCR in all experimental groups, for normal mice it was negative. The results indicated that the systemic disseminated leukemia model was established successfully by caudal vein injection 8×105/mouse Kasumi-1 cells in the three experimental groups.